PROJECT SUMMARY/ABSTRACT Candida albicans, the primary causative agent of fungal vaginitis, will affect 75% of all women of reproductive age at least once in their lifetime. Long believed to result from immunodeficiency, a growing body of evidence strongly suggests that vaginitis is now considered to be an immunopathology, in which the host response actually drives disease symptoms. This fact, coupled with static activity of commonly deployed antifungals, high incidence of chronic infection, and emerging antifungal resistance necessitates a shift in treatment strategy. Administration of azole antifungals, both systemically and topically, remains the go-to treatment for vaginal candidiasis. While vaginal fungal burden is reduced in a majority of treated women, it can take days to a week for ablation of symptoms. Therefore, inhibiting the robust host inflammatory response may serve as novel adjunctive therapy to break the cycle of inflammation and resolve disease symptoms in a timely fashion. Guided by strong preliminary data, we have identified the FDA-approved sulfonylureas (commonly used as anti-hyperglycemic agents) as potential therapeutic agents against symptomatic vulvovaginal candidiasis. Aside from anti-hyperglycemic effects, some sulfonylureas also possess anti-inflammatory activity by inhibiting assembly of the inflammasome complex. Therefore, the objective of this proposal is to systematically evaluate the anti-inflammasome activity exhibited by various commonly used sulfonylureas in a variety of cell types and to determine their efficacy in a murine model of vaginitis. These aims will test our central hypothesis that sulfonylureas may serve as repurposed agents effective at reducing vaginitis immunopathology. Under the first aim, a systematic approach to assess the ability of sulfonylureas to prevent expression of inflammasome effectors (IL-1?, IL-18) in wild-type macrophages or those deficient in specific inflammasome components challenged with C. albicans will reveal inflammasome-specificity of this drug class. Secondary assays will evaluate anti-inflammatory effects of sulfonylureas against neutrophils, vaginal epithelium, and primary mononuclear cells. Toxicity at doses required for anti-inflammatory activity will also be addressed. The second aim will focus on defining optimal doses, delivery methods, and timing of administration for sulfonylureas (identified in prior assays) to inhibit immunopathology (neutrophil influx, inflammatory cytokines) in a mouse model of vaginal candidiasis. Experiments to assess co-therapeutic potential of sulfonylureas with azoles will also be conducted. The outcomes of this project will provide foundational information for the utility of sulfonylureas to treat vaginitis and also raise questions regarding off-target effects of commonly used anti- hyperglycemics in the diabetic population. Furthermore, The objectives of this R21 proposal fulfill the NIAID's mission to identify novel therapeutic approaches to combat fungal disease and challenges current treatment paradigms.